New insights into azelaic acid-induced resistance against Alternaria Solani in tomato plants.

BACKGROUND: The effect of azelaic acid (Aza) on the response of tomato plants to Alternaria solani was investigated in this study. After being treated with Aza, tomato plants were infected with A. solani, and their antioxidant, biochemical, and molecular responses were analyzed. RESULTS: The results demonstrated that H2O2 and MDA accumulation increased in control plants after pathogen infection. Aza-treated plants exhibited a remarkable rise in peroxidase (POD) and catalase (CAT) activities during the initial stages of A. solani infection. Gene expression analysis revealed that both Aza treatment and pathogen infection altered the expression patterns of the SlNPR1, SlERF2, SlPR1, and SlPDF1.2 genes. The expression of SlPDF1.2, a marker gene for the jasmonic acid/ethylene (JA/ET) signaling pathway, showed a remarkable increase of 4.2-fold upon pathogen infection. In contrast, for the SlNPR1, a key gene in salicylic acid (SA) pathway, this increased expression was recorded with a delay at 96 hpi. Also, the phytohormone analysis showed significantly increased SA accumulation in plant tissues with disease development. It was also revealed that tissue accumulation of JA in Aza-treated plants was increased following pathogen infection, while it was not increased in plants without pathogen inoculation. CONCLUSION: The results suggest that the resistance induced by Aza is mainly a result of modulations in both SA and JA pathways following complex antioxidant and molecular defense responses in tomato plants during A. solani infection. These findings provide novel information regarding inducing mechanisms of azelaic acid which would add to the current body of knowledge of SAR induction in plants as result of Aza application.

Beauveria bassiana rewires molecular mechanisms related to growth and defense in tomato.

Plant roots can exploit beneficial associations with soil-inhabiting microbes, promoting growth and expanding the immune capacity of the host plant. In this work, we aimed to provide new information on changes occurring in tomato interacting with the beneficial fungus Beauveria bassiana. The tomato leaf proteome revealed perturbed molecular pathways during the establishment of the plant-fungus relationship. In the early stages of colonization (5-7 d), proteins related to defense responses to the fungus were down-regulated and proteins related to calcium transport were up-regulated. At later time points (12-19 d after colonization), up-regulation of molecular pathways linked to protein/amino acid turnover and to biosynthesis of energy compounds suggests beneficial interaction enhancing plant growth and development. At the later stage, the profile of leaf hormones and related compounds was also investigated, highlighting up-regulation of those related to plant growth and defense. Finally, B. bassiana colonization was found to improve plant resistance to Botrytis cinerea, impacting plant oxidative damage. Overall, our findings further expand current knowledge on the possible mechanisms underlying the beneficial role of B. bassiana in tomato plants.

The AP2/ERF transcription factor ORA59 regulates ethylene-induced phytoalexin synthesis through modulation of an acyltransferase gene expression.

The gaseous ethylene (ET) and the oxylipin-derived jasmonic acid (JA) in plants jointly regulate an arsenal of pathogen responsive genes involved in defending against necrotrophic pathogens. The APETALA2 (AP2)/ETHYLENE RESPONSE FACTOR (ERF) transcription factor ORA59 is a major positive regulator of the ET/JA-mediated defense pathway in Arabidopsis thaliana. The Arabidopsis agmatine coumaroyltransferase (AtACT) catalyzes the formation of hydroxycinnamic acid amides (HCAAs) which are effective toxic antimicrobial substances known as phytoalexins and play an important role in plant defense response. However, induction and regulation of AtACT gene expression and HCAAs synthesis in plants remain less understood. Through gene coexpression network analysis, we identified a list of GCC-box cis-element containing genes that were coexpressed with ORA59 under diverse biotic stress conditions and might be potential downstream targets of this AP2/ERF-domain transcription factor. Particularly, ORA59 directly binds to AtACT gene promoter via the GCC-boxes and activates AtACT gene expression. The ET precursor 1-aminocyclopropane-1-carboxylic acid (ACC)-treatment significantly induces AtACT gene expression. Both ORA59 and members of the class II TGA transcription factors are indispensable for ACC-induced AtACT expression. Interestingly, the expression of AtACT is also subject to the signaling crosstalk of the salicylic acid- and ET/JA-mediated defense response pathways. In addition, we found that genes of the phenylpropanoid metabolism pathway were specifically induced by Botrytis cinerea. Taking together, these evidence suggest that the ET/JA signaling pathway activate the expression of AtACT to increase antimicrobial HCAAs production through the transcription factor ORA59 in response to the infection of necrotrophic plant pathogens.

AUXIN RESPONSE FACTOR 16 (StARF16) regulates defense gene StNPR1 upon infection with necrotrophic pathogen in potato.

KEY MESSAGE: We demonstrate a new regulatory mechanism in the jasmonic acid (JA) and salicylic acid (SA) mediated crosstalk in potato defense response, wherein, miR160 target StARF16 (a gene involved in growth and development) binds to the promoter of StNPR1 (a defense gene) and negatively regulates its expression to suppress the SA pathway. Overall, our study establishes the importance of StARF16 in regulation of StNPR1 during JA mediated defense response upon necrotrophic pathogen interaction. Plants employ antagonistic crosstalk between salicylic acid (SA) and jasmonic acid (JA) to effectively defend them from pathogens. During biotrophic pathogen attack, SA pathway activates and suppresses the JA pathway via NONEXPRESSOR OF PATHOGENESIS-RELATED GENES 1 (NPR1). However, upon necrotrophic pathogen attack, how JA-mediated defense response suppresses the SA pathway, is still not well-understood. Recently StARF10 (AUXIN RESPONSE FACTOR), a miR160 target, has been shown to regulate SA and binds to the promoter of StGH3.6 (GRETCHEN HAGEN3), a gene proposed to maintain the balance between the free SA and auxin in plants. In the current study, we investigated the role of StARF16 (a miR160 target) in the regulation of the defense gene StNPR1 in potato upon activation of the JA pathway. We observed that a negative correlation exists between StNPR1 and StARF16 upon infection with the pathogen. The results were further confirmed through the exogenous application of SA and JA. Using yeast one-hybrid assay, we demonstrated that StARF16 binds to the StNPR1 promoter through putative ARF binding sites. Additionally, through protoplast transfection and chromatin immunoprecipitation experiments, we showed that StARF16 could bind to the StNPR1 promoter and regulate its expression. Co-transfection assays using promoter deletion constructs established that ARF binding sites are present in the 2.6 kb sequence upstream to the StNPR1 gene and play a key role in its regulation during infection. In summary, we demonstrate the importance of StARF16 in the regulation of StNPR1, and thus SA pathway, during JA-mediated defense response upon necrotrophic pathogen interaction.

Interaction of drought- and pathogen-induced mortality in Norway spruce and Scots pine.

Pathogenic diseases frequently occur in drought-stressed trees. However, their contribution to the process of drought-induced mortality is poorly understood. We combined drought and stem inoculation treatments to study the physiological processes leading to drought-induced mortality in Norway spruce (Picea abies) and Scots pine (Pinus sylvestris) saplings infected with Heterobasidion annosum s.s. We analysed the saplings' water status, gas exchange, nonstructural carbohydrates (NSCs) and defence responses, and how they related to mortality. Saplings were followed for two growing seasons, including an artificially induced 3-month dormancy period. The combined drought and pathogen treatment significantly increased spruce mortality; however, no interaction between these stressors was observed in pine, although individually each stressor caused mortality. Our results suggest that pathogen infection decreased carbon reserves in spruce, reducing the capacity of saplings to cope with drought, resulting in increased mortality rates. Defoliation, relative water content and the starch concentration of needles were predictors of mortality in both species under drought and pathogen infection. Infection and drought stress create conflicting needs for carbon to compartmentalize the pathogen and to avoid turgor loss, respectively. Heterobasidion annosum reduces the functional sapwood area and shifts NSC allocation patterns, reducing the capacity of trees to cope with drought.

Floral Microbes Suppress Growth of Monilinia laxa with Minimal Effects on Honey Bee Feeding.

Management of Monilinia laxa, the causal agent of brown rot blossom blight in almond (Prunus dulcis), relies heavily on the use of chemical fungicides during bloom. However, chemical fungicides can have nontarget effects on beneficial arthropods, including pollinators, and select for resistance in the pathogen of concern. Almond yield is heavily reliant on successful pollination by healthy honey bees (Apis mellifera); thus, identifying sustainable, effective, and pollinator-friendly control methods for blossom blight during bloom is desirable. Flower-inhabiting microbes could provide a natural, sustainable form of biocontrol for M. laxa, while potentially minimizing costly nontarget effects on almond pollinators and the services they provide. As pollinators are sensitive to floral microbes and their associated taste and scent cues, assessing effects of prospective biocontrol species on pollinator attraction is also necessary. Here, our objective was to isolate and identify potential biocontrol microbes from an array of agricultural and natural flowering hosts and test their efficacy in suppressing M. laxa growth in culture. Out of an initial 287 bacterial and fungal isolates identified, 56 were screened using a dual culture plate assay. Most strains reduced M. laxa growth in vitro. Ten particularly effective candidate microbes were further screened for their effect on honey bee feeding. Of the 10, nine were found to both strongly suppress M. laxa growth in culture and not reduce honey bee feeding. These promising results suggest a number of strong candidates for augmentative microbial biocontrol of brown rot blossom blight in almond with potentially minimal effects on honey bee pollination.

The transcription factor WRKY75 positively regulates jasmonate-mediated plant defense to necrotrophic fungal pathogens.

Necrotrophic fungi cause devastating diseases in both horticultural and agronomic crops, but our understanding of plant defense responses against these pathogens is still limited. In this study, we demonstrated that WRKY75 positively regulates jasmonate (JA)-mediated plant defense against necrotrophic fungal pathogens Botrytis cinerea and Alternaria brassicicola, and also affects the sensitivity of plants to JA-inhibited seed germination and root growth. Quantitative analysis indicated that several JA-associated genes, such as OCTADECANOID-RESPONSIVE ARABIDOPSIS (ORA59) and PLANT DEFENSIN 1.2A (PDF1.2), were significantly reduced in expression in wrky75 mutants, and enhanced in WRKY75 overexpressing transgenic plants. Immunoprecipitation assays revealed that WRKY75 directly binds to the promoter of ORA59 and represses itstranscription. In vivo and in vitro experiments suggested that WRKY75 interacts with several JASMONATE ZIM-domain proteins, repressors of the JA signaling pathway. We determined that JASMONATE-ZIM-DOMAIN PROTEIN 8 (JAZ8) represses the transcriptional function of WRKY75, thereby attenuating the expression of its regulation. Overexpression of JAZ8 repressed plant defense responses to B. cinerea. Our study provides evidence that WRKY75 functions as a critical component of the JA-mediated signaling pathway to positively regulate Arabidopsis defense responses to necrotrophic pathogens.

Prior exposure of Arabidopsis seedlings to mechanical stress heightens jasmonic acid-mediated defense against necrotrophic pathogens.

BACKGROUND: Prolonged mechanical stress (MS) causes thigmomorphogenesis, a stress acclimation response associated with increased disease resistance. What remains unclear is if; 1) plants pre-exposed to a short period of repetitive MS can prime defence responses upon subsequent challenge with necrotrophic pathogens, 2) MS mediates plant immunity via jasmonic acid (JA) signalling, and 3) a short period of repetitive MS can cause long-term changes in gene expression resembling a stress-induced memory. To address these points, 10-days old juvenile Arabidopsis seedlings were mechanically stressed for 7-days using a soft brush and subsequently challenged with the necrotrophic pathogens, Alternaria brassicicola, and Botrytis cinerea. Here we assessed how MS impacted structural cell wall appositions, disease symptoms and altered gene expression in response to infection. RESULTS: The MS-treated plants exhibited enhanced cell wall appositions and jasmonic acid (JA) accumulation that correlated with a reduction in disease progression compared to unstressed plants. The expression of genes involved in JA signalling, callose deposition, peroxidase and phytoalexin biosynthesis and reactive oxygen species detoxification were hyper-induced 4-days post-infection in MS-treated plants. The loss-of-function in JA signalling mediated by the JA-insensitive coronatine-insensitive 1 (coi1) mutant impaired the hyper-induction of defense gene expression and promoted pathogen proliferation in MS-treated plants subject to infection. The basal expression level of PATHOGENESIS-RELATED GENE 1 and PLANT DEFENSIN 1.2 defense marker genes were constitutively upregulated in rosette leaves for 5-days post-MS, as well as in naïve cauline leaves that differentiated from the inflorescence meristem well after ceasing MS. CONCLUSION: This study reveals that exposure of juvenile Arabidopsis plants to a short repetitive period of MS can alter gene expression and prime plant resistance upon subsequent challenge with necrotrophic pathogens via the JA-mediated COI1 signalling pathway. MS may facilitate a stress-induced memory to modulate the plant's response to future stress encounters. These data advance our understanding of how MS primes plant immunity against necrotrophic pathogens and how that could be utilised in sustainable agricultural practices.

Chlorophyll fluorescence imaging for monitoring effects of Heterobasidion parviporum small secreted protein induced cell death and in planta defense gene expression.

Heterobasidion parviporum Niemelä & Korhonen is a necrotrophic fungal pathogen of Norway spruce (Picea abies). The H. parviporum genome encodes numerous necrotrophic small secreted proteins (SSP) which might be important for promoting and sustaining the disease development. However, their transcriptional dynamics and plant defense response during infection are largely unknown. In this study, we identified a necrotrophic SSP named HpSSP35.8 and its coding gene was highly expressed in the pre-symptomatic phase of the host (Norway spruce) infection. We explored the impact of HpSSP35.8 on non-host Nicotiana benthamiana using Agrobacterium-mediated transient expression system under visible spectrum RGB imaging and chlorophyll fluorescence imaging. The results showed that HpSSP35.8 triggered a form of SSP-associated programmed cell death, accompanied by a decrease in the plant photosynthetic activity. Defense-related genes including WRKY12, ethylene response factor (ERF1α) and a chitinase gene PR4 were up-regulated in both HpSSP35.8-N. benthamiana interaction and H. parviporum-Norway spruce pathosystem. This study also highlighted the potential to use the chlorophyll fluorescence imaging approach to monitor both the indirect effects of SSP and also for the selection of other potential effector-like protein candidates.

Marker development, saturation mapping, and high-resolution mapping of the Septoria nodorum blotch susceptibility gene Snn3-B1 in wheat.

Septoria nodorum blotch (SNB), caused by Parastagonospora nodorum, is a severe foliar and glume disease on durum and common wheat. Pathogen-produced necrotrophic effectors (NEs) are the major determinants for SNB on leaves. One such NE is SnTox3, which evokes programmed cell death and leads to disease when recognized by the wheat Snn3-B1 gene. Here, we developed saturated genetic linkage maps of the Snn3-B1 region using two F2 populations derived from the SnTox3-sensitive line Sumai 3 crossed with different SnTox3-insensitive lines. Markers were identified and/or developed from various resources including previously mapped simple sequence repeats, bin-mapped expressed sequence tags, single nucleotide polymorphisms, and whole genome survey sequences. Subsequent high-resolution mapping of the Snn3-B1 locus in 5600 gametes delineated the gene to a 1.5 cM interval. Analysis of micro-colinearity of the Snn3-B1 region indicated that it was highly disrupted compared to rice and Brachypodium distachyon. The screening of a collection of durum and common wheat cultivars with tightly linked markers indicated they are not diagnostic for the presence of Snn3-B1, but can be useful for marker-assisted selection if the SnTox3 reactions of lines are first determined. Finally, we developed an ethyl methanesulfonate-induced mutant population of Sumai 3 where the screening of 408 M2 families led to the identification of 17 SnTox3-insensitive mutants. These mutants along with the markers and high-resolution map developed in this research provide a strong foundation for the map-based cloning of Snn3-B1, which will broaden our understanding of the wheat-P. nodorum system and plant-necrotrophic pathogen interactions in general.

Nitrogen and Silicon Contribute to Wheat Defense's to Pyrenophora tritici-repentis, but in an Independent Manner.

Nitrogen (N) and silicon (Si) are mineral elements that have shown a reduction in the damage caused by tan spot (Pyrenophora tritici-repentis (Ptr)) in wheat. However, the effects of these elements were studied separately, and the N and Si interaction effect on wheat resistance to tan spot remains elusive. Histocytological and biochemical defense responses against Ptr in wheat leaves treated with Si (+Si) at low (LN) and high N (HN) inputs were investigated. Soil amendment with Si reduced the tan spot severity in 18% due to the increase in the leaf Si concentration (around 30%), but it was affected by the N level used. The superoxide dismutase (SOD) activity was higher in +Si plants and inoculated with Ptr, leading to early and higher H2O2 and callose accumulation in wheat leaf. Interestedly, phenylalanine ammonia-lyase (PAL) activity was induced by the Si supplying, being negatively affected by the HN rate. Meanwhile, catalase (CAT), and peroxidase (POX) activities showed differential response patterns according to the Si and N rates used. Tan spot severity was reduced by both elements, but their interaction does not evidence synergic effects in this disease's control. Wheat plants from -Si and HN and +Si and LN treatments recorded lower tan spot severity.

Colonization by orchid mycorrhizal fungi primes induced systemic resistance against necrotrophic pathogen.

Orchids and arbuscular mycorrhiza (AM) plants evolved independently and have different structures and fungal partners, but they both facilitate nutrient uptake. Orchid mycorrhiza (OM) supports orchid seed germination, but unlike AM, its role in disease resistance of mature plants is largely unknown. Here, we examined whether OM induces systemic disease resistance against a necrotrophic pathogen in a similar fashion to AM. We investigated the priming effect of mycorrhizal fungi inoculation on resistance of a terrestrial orchid, Bletilla striata, to soft rot caused by Dickeya fangzhongdai. We found that root colonization by a compatible OM fungus primed B. striata seedlings and induced systemic resistance against the infection. Transcriptome analysis showed that priming was mediated by the downregulation of jasmonate and ethylene pathways and that these pathways are upregulated once infection occurs. Comparison with the reported transcriptome of AM fungus-colonized rice leaves revealed similar mechanisms in B. striata and in rice. These findings highlight a novel aspect of commonality between OM and AM plants in terms of induced systemic resistance.

The Bacterial Volatile Organic Compound N,N-Dimethylhexadecylamine Induces Long-Lasting Developmental and Immune Responses throughout the Life Cycle of Arabidopsis thaliana.

N,N-dimethylhexadecylamine (DMHDA) is a bacterial volatile organic compound that affects plant growth and morphogenesis and is considered a cross-kingdom signal molecule. Its bioactivity involves crosstalk with the cytokinin and jasmonic acid (JA) pathways to control stem cell niches and induce iron deficiency adaptation and plant defense. In this study, through genetic analysis, we show that the DMHDA-JA-Ethylene (ET) relations determine the magnitude of the defensive response mounted during the infestation of Arabidopsis plants by the pathogenic fungus Botrytis cinerea. The Arabidopsis mutants defective in the JA receptor CORONATINE INSENSITIVE 1 (coi1-1) showed a more severe infestation when compared to wild-type plants (Col-0) that were partially restored by DMHDA supplements. Moreover, the oversensitivity manifested by ETHYLENE INSENSITIVE 2 (ein2) by B. cinerea infestation could not be reverted by the volatile, suggesting a role for this gene in DMHDA reinforcement of immunity. Growth of Col-0 plants was inhibited by DMHDA, but ein2 did not. Noteworthy, Arabidopsis seeds treated with DMHDA produced more vigorous plants throughout their life cycle. These data are supportive of a scenario where plant perception of a bacterial volatile influences the resistance to a fungal phytopathogen while modulating plant growth.

Plant defense: ARR11 response regulator as a potential player in Arabidopsis.

Plant growth and response to environmental cues are largely driven by hormones. Salicylic acid (SA)- and jasmonic acid (JA)-mediated defenses have been shown to be effective against different types of attackers. SA-mediated defense is mainly effective against biotrophic pathogens and phloem-feeding insects, whereas JA-mediated defense is effective against necrotrophic pathogens and tissue-damaging insects. Cytokinins (CKs) are classic growth hormones that have also emerged as plant immunity modulators. Evidence pointed out that CKs contribute to the defense responses mediated by SA and JA, acting as hormone modulators of the SA/JA signaling backbone. Recently, we identified in Arabidopsis a type-B response regulator 11 (ARR 11) involved in cytokinin-mediated responses as a novel regulator of the SA/JA cross-talk. Here we investigated plant fitness and resistance against the fungal necrotrophic pathogen Botrytis cinerea in Arabidopsis wild-type Col-8 and defective arr11 mutant following SA, JA, CK single or combined treatment. Our results demonstrated that the CK and SA/JA/CK combination has a positive outcome on plant fitness in both Arabidopsis Col-8 and arr11 mutant,. The triple hormone treatment is efficient in increasing resistance to B. cinerea in Col-8 and this effect is stronger in arr11 mutant. The results will provide not only new background knowledge, corroborating the role of ARR11 in plant-defense related processes, but also new potential opportunities for alternative ways of protecting plants from fungal diseases.

Volatile Metabolism of Wine Grape Trincadeira: Impact of Infection with Botrytis cinerea.

The aroma of grapes is cultivar dependent and is influenced by terroir, vineyard practices, and abiotic and biotic stresses. Trincadeira is a non-aromatic variety associated with low phenolic content and high sugar and organic acid levels. This cultivar, widely used in Portuguese wines, presents high susceptibility to Botrytis cinerea. This work aimed to characterise the volatile profile of Trincadeira grapes and how it changes under infection with B. cinerea. Thirty-six volatile organic compounds were identified, from different functional groups, namely alcohols, ester acetates, fatty acid esters, fatty acids, aldehydes, and products of the lipoxygenase pathway. Both free and glycosidic volatile organic compounds were analysed by Gas Chromatography and Gas Chromatography coupled to Mass Spectrometry for component quantification and identification, respectively. A multivariance analysis showed a clear discrimination between healthy and infected grapes with 2-trans-hexenal and isoamyl-acetate among the compounds identified as negative and positive markers of infection, respectively. Ester acetates such as 2-phenylethyl acetate, isoamyl acetate, and 2-methylbutyl acetate were present in higher contents in infected samples, whereas the contents of several fatty acid esters, such as ethyl decanoate and ethyl dodecanoate, decreased. These data were integrated with quantitative PCR data regarding genes involved in volatile metabolism and showed up-regulation of a gene coding for Hydroperoxide Lyase 2 in infected grapes. Altogether, these changes in volatile metabolism indicate an impact on the grape quality and may be related to defence against B. cinerea. The presence/absence of specific compounds might be used as infection biomarkers in the assessment of Trincadeira grapes' quality.

Tenuazonic Acid-Triggered Cell Death Is the Essential Prerequisite for Alternaria alternata (Fr.) Keissler to Infect Successfully Host Ageratina adenophora.

The necrotrophic fungus Alternaria alternata contains different pathotypes that produce different mycotoxins. The pathotype Ageratina adenophora secretes the non-host-selective toxin tenuazonic acid (TeA), which can cause necrosis in many plants. Although TeA is thought to be a central virulence factor of the A. adenophora pathotype, the precise role of TeA in different stages of host infection by pathogens remains unclear. Here, an A. alternata wild-type and the toxin-deficient mutant ΔHP001 with a 75% reduction in TeA production were used. It was observed that wild-type pathogens could induce the reactive oxygen species (ROS) bursts in host leaves and killed photosynthetic cells before invading hyphae. The ROS interceptor catalase remarkably inhibited hyphal penetration and invasive hyphal growth and expansion in infected leaves and suppressed necrotic leaf lesion. This suggests that the production of ROS is critical for pathogen invasion and proliferation and disease symptom formation during infection. It was found that the mutant pathogens did not cause the formation of ROS and cell death in host leaves, showing an almost complete loss of disease susceptibility. In addition, the lack of TeA resulted in a significant reduction in the ability of the pathogen to penetrate invasive hyphal growth and spread. The addition of exogenous TeA, AAL-toxin, and bentazone to the mutant ΔHP001 pathogens during inoculation resulted in a significant restoration of pathogenicity by increasing the level of cell death, frequency of hyphal penetration, and extent of invasive hyphal spread. Our results suggest that cell death triggered by TeA is the essential requirement for successful colonization and disease development in host leaves during infection with A. adenophora pathogens.

Ciboria carunculoides Suppresses Mulberry Immune Responses Through Regulation of Salicylic Acid Signaling.

Ciboria carunculoides is the dominant causal agent of mulberry sclerotial disease, and it is a necrotrophic fungal pathogen with a narrow host range that causes devastating diseases in mulberry fruit. However, little is known about the interaction between C. carunculoides and mulberry. Here, our transcriptome sequencing results showed that the transcription of genes in the secondary metabolism and defense-related hormone pathways were significantly altered in infected mulberry fruit. Due to the antimicrobial properties of proanthocyanidins (PAs), the activation of PA biosynthetic pathways contributes to defense against pathogens. Salicylic acid (SA) and jasmonic acid (JA) are major plant defense hormones. However, SA signaling and JA signaling are antagonistic to each other. Our results showed that SA signaling was activated, while JA signaling was inhibited, in mulberry fruit infected with C. carunculoides. Yet SA mediated responses are double-edged sword against necrotrophic pathogens, as SA not only activates systemic acquired resistance (SAR) but also suppresses JA signaling. We also show here that the small secreted protein CcSSP1 of C. carunculoides activates SA signaling by targeting pathogenesis-related protein 1 (PR1). These findings reveal that the infection strategy of C. carunculoides functions by regulating SA signaling to inhibit host defense responses.

Plant surface metabolites as potent antifungal agents.

Triunsaturated fatty acids are substrates for the synthesis of the defense hormone jasmonate which plays roles in resistance to numerous fungal pathogens. However, relatively little is known about other potential roles of di-unsaturated and triunsaturated fatty acids in resistance to fungal pathogens - in particular those that can attack plants at the seedling stage. We examined the roles of polyunsaturated fatty acids (PUFAs) in Arabidopsis thaliana during attack by the necrotrophic pathogen, Botrytis cinerea. We found that PUFA-deficient Arabidopsis mutants (fad2-1, fad2-3 and fad3-2 fad7-2 fad8 [fad trip]) displayed an unexpectedly strong resistance to B. cinerea at the cotyledon stage. Preliminary analyses revealed no changes in the expression of defense genes, however cuticle permeability defects were detected in both fad2-1 and fad trip mutants. Analysis of B. cinerea development on the surface of cotyledons revealed arrested hyphal growth on fad2-3 and fad trip mutants and 28% reduction in fungal adhesion on fad2-3 cotyledons. Surface metabolite analysis from the cotyledons of PUFA mutants led to the identification of 7-methylsulfonylheptyl glucosinolate (7MSOHG), which over-accumulated on the plant surface. We linked the appearance of 7MSOHG to defects in cuticle composition and permeability of mutants and show that its appearance correlates with resistance to B. cinerea.

The study of hormonal metabolism of Trincadeira and Syrah cultivars indicates new roles of salicylic acid, jasmonates, ABA and IAA during grape ripening and upon infection with Botrytis cinerea.

Hormones play an important role in fruit ripening and in response to biotic stress. Nevertheless, analyses of hormonal profiling during plant development and defense are scarce. In this work, changes in hormonal metabolism in grapevine (Vitis vinifera) were compared between a susceptible (Trincadeira) and a tolerant (Syrah) variety during grape ripening and upon infection with Botrytis cinerea. Infection of grapes with the necrotrophic pathogen Botrytis cinerea leads to significant economic losses worldwide. Peppercorn-sized fruits were infected in the field and mock-treated and infected berries were collected at green, veraison and harvest stages for hormone analysis and targeted qPCR analysis of genes involved in hormonal metabolism and signaling. Results indicate a substantial reprogramming of hormonal metabolism during grape ripening and in response to fungal attack. Syrah and Trincadeira presented differences in the metabolism of abscisic acid (ABA), indole-3-acetic acid (IAA) and jasmonates during grape ripening that may be connected to fruit quality. On the other hand, high basal levels of salicylic acid (SA), jasmonates and IAA at an early stage of ripening, together with activated SA, jasmonates and IAA signaling, likely enable a fast defense response leading to grape resistance/ tolerance towards B. cinerea. The balance among the different phytohormones seems to depend on the ripening stage and on the intra-specific genetic background and may be fundamental in providing resistance or susceptibility. In addition, this study indicated the involvement of SA and IAA in defense against necrotrophic pathogens and gains insights into possible strategies for conventional breeding and/or gene editing aiming at improving grape quality and grape resistance against Botrytis cinerea.

Paenibacillus lentimorbus induces autophagy for protecting tomato from Sclerotium rolfsii infection.

During biotic stress, plants use several mechanisms to protect themselves that include the production of reactive oxygen species (ROS), induction of pathogenesis-related proteins and cell death. Some plant growth promoting rhizobacteria (PGPR) are known to act as bio-control agents that protect crops against pathogens. The biocontrol activity of PGPR Paenibacillus lentimorbus (B-30488) against Sclerotium rolfsii showed previously where several defense-related genes were upregulated with ROS induction in tomato. We further evaluate the other possibility, i.e. role of autophagy in enhancing defense in tomato using PGPR. Confocal microscopy revealed the presence of an acidotropic dye Mono Dansyl Cadaverine (MDC) stained autophagosomes in B-30488 treated healthy and infected plants. These autophagosomes almost disappeared in plants treated with an autophagy inhibitor chloroquine. The results were also confirmed by ultrastructural analysis of leaf tissues using transmission electron microscopy. Enhanced expression of autophagy-related genes was also monitored in B-30488 primed fungal infected tissues as compared to control by qRT-PCR. Results of ROS accumulation, fluorescence, confocal and transmission electron microscopy and gene expression analysis revealed induction of autophagy using B-30488 as a biocontrol agent suggesting a role in enhancing disease resistance in tomato. Overall, the present study indicated a role of B-30488 as a biocontrol in enhancing disease resistance in tomato and also assists a better understanding of fungal pathogenesis that is expected to be useful in developing new strategies for disease control.